Prolyl hydroxylase PHD3 activates oxygen-dependent protein aggregation

The HIF prolyl hydroxylases (PHDs/EGLNs) are central regulators of the molecular responses to oxygen availability. One isoform, PHD3, is expressed in response to hypoxia and causes apoptosis in oxygenated conditions in neural cells. Here we show that PHD3 forms subcellular aggregates in an oxygen-dependent manner. The aggregation of PHD3 was seen under normoxia and was strongly reduced under hypoxia or by the inactivation of the PHD3 hydroxylase activity. The PHD3 aggregates were dependent on microtubular integrity and contained components of the 26S proteasome, chaperones, and ubiquitin, thus demonstrating features that are characteristic for aggresome-like structures. Forced expression of the active PHD3 induced the aggregation of proteasomal components and activated apoptosis under normoxia in HeLa cells. The apoptosis was seen in cells prone to PHD3 aggregation and the PHD3 aggregation preceded apoptosis. The data demonstrates the cellular oxygen sensor PHD3 as a regulator of protein aggregation in response to varying oxygen availability

Surgical robotics beyond enhanced dexterity instrumentation: a survey of machine learning techniques and their role in intelligent and autonomous surgical actions

PURPOSE: Advances in technology and computing play an increasingly important role in the evolution of modern surgical techniques and paradigms. This article reviews the current role of machine learning (ML) techniques in the context of surgery with a focus on surgical robotics (SR). Also, we provide a perspective on the future possibilities for enhancing the effectiveness of procedures by integrating ML in the operating room. METHODS: The review is focused on ML techniques directly applied to surgery, surgical robotics, surgical training and assessment. The widespread use of ML methods in diagnosis and medical image computing is beyond the scope of the review. Searches were performed on PubMed and IEEE Explore using combinations of keywords: ML, surgery, robotics, surgical and medical robotics, skill learning, skill analysis and learning to perceive. RESULTS: Studies making use of ML methods in the context of surgery are increasingly being reported. In particular, there is an increasing interest in using ML for developing tools to understand and model surgical skill and competence or to extract surgical workflow. Many researchers begin to integrate this understanding into the control of recent surgical robots and devices. CONCLUSION: ML is an expanding field. It is popular as it allows efficient processing of vast amounts of data for interpreting and real-time decision making. Already widely used in imaging and diagnosis, it is believed that ML will also play an important role in surgery and interventional treatments. In particular, ML could become a game changer into the conception of cognitive surgical robots. Such robots endowed with cognitive skills would assist the surgical team also on a cognitive level, such as possibly lowering the mental load of the team. For example, ML could help extracting surgical skill, learned through demonstration by human experts, and could transfer this to robotic skills. Such intelligent surgical assistance would significantly surpass the state of the art in surgical robotics. Current devices possess no intelligence whatsoever and are merely advanced and expensive instruments

The Myth of the Stupid Believer: The Negative Religiousness–IQ Nexus is Not on General Intelligence (g) and is Likely a Product of the Relations Between IQ and Autism Spectrum Traits

Numerous studies have found a negative relationship between religiousness and IQ. It is in the region of −0.2, according to meta-analyses. The reasons for this relationship are, however, unknown. It has been suggested that higher intelligence leads to greater attraction to science, or that it helps to override evolved cognitive dispositions such as for religiousness. Either way, such explanations assume that the religion–IQ nexus is on general intelligence (g), rather than some subset of specialized cognitive abilities. In other words, they assume it is a Jensen efect. Two large datasets comparing groups with diferent levels of religiousness show that their IQ diferences are not on g and must, therefore, be attributed to specialized abilities. An analysis of the specialized abilities on which the religious and non-religious groups difer reveals no clear pattern. We cautiously suggest that this may be explicable in terms of autism spectrum disorder traits among people with high IQ scores, because such traits are negatively associated with religiousness

Upside-down fluxes Down Under: CO2 net sink in winter and net source in summer in a temperate evergreen broadleaf forest

Predicting the seasonal dynamics of ecosystem carbon fluxes is challenging in broadleaved evergreen forests because of their moderate climates and subtle changes in canopy phenology. We assessed the climatic and biotic drivers of the seasonality of net ecosystem–atmosphere CO2 exchange (NEE) of a eucalyptus-dominated forest near Sydney, Australia, using the eddy covariance method. The climate is characterised by a mean annual precipitation of 800mm and a mean annual temperature of 18°C, hot summers and mild winters, with highly variable precipitation. In the 4-year study, the ecosystem was a sink each year (−225gCm−2yr−1 on average, with a standard deviation of 108gCm−2yr−1); inter-annual variations were not related to meteorological conditions. Daily net C uptake was always detected during the cooler, drier winter months (June through August), while net C loss occurred during the warmer, wetter summer months (December through February). Gross primary productivity (GPP) seasonality was low, despite longer days with higher light intensity in summer, because vapour pressure deficit (D) and air temperature (Ta) restricted surface conductance during summer while winter temperatures were still high enough to support photosynthesis. Maximum GPP during ideal environmental conditions was significantly correlated with remotely sensed enhanced vegetation index (EVI; r2 = 0.46) and with canopy leaf area index (LAI; r2= 0.29), which increased rapidly after mid-summer rainfall events. Ecosystem respiration (ER) was highest during summer in wet soils and lowest during winter months. ER had larger seasonal amplitude compared to GPP, and therefore drove the seasonal variation of NEE. Because summer carbon uptake may become increasingly limited by atmospheric demand and high temperature, and because ecosystem respiration could be enhanced by rising temperatures, our results suggest the potential for large-scale seasonal shifts in NEE in sclerophyll vegetation under climate change.The Australian Education Investment Fund, Australian Terrestrial Ecosystem Research Network, Australian Research Council and Hawkesbury Institute for the Environment at Western Sydney University supported this work. We thank Jason Beringer, Helen Cleugh, Ray Leuning and Eva van Gorsel for advice and support. Senani Karunaratne provided soil classification details

PHD3 regulates differentiation, tumour growth and angiogenesis in pancreatic cancer

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The Function of Hypoxia-Inducible Factor (HIF) Is Independent of the Endoplasmic Reticulum Protein OS-9

The protein “amplified in osteosarcoma-9” (OS-9) has been shown previously to interact with the prolyl hydroxylases PHD2 and PHD3. These enzymes initiate oxygen-dependent degradation of the α-subunit of hypoxia-inducible factor (HIF), a transcription factor that adapts cells to insufficient oxygen supply (hypoxia). A new model has been proposed where OS-9 triggers PHD dependent degradation of HIF-α. It was the aim of our study to define the molecular mode of action of OS-9 in the regulation of PHD and HIF activity. Although initial co-immunoprecipitation experiments confirmed physical interaction between OS-9 and PHD2, neither overexpression nor lentiviral inhibition of OS-9 expression affected HIF regulation. Subcellular localization experiments revealed a distinct reticular staining pattern for OS-9 while PHD2 was mainly localized in the cytoplasm. Further cell fractionation experiments and glycosylation tests indicated that OS-9 is a luminal ER protein. In vivo protein interaction analysis by fluorescence resonance energy transfer (FRET) showed no significant physical interaction of overexpressed PHD2-CFP and OS-9-YFP. We conclude that OS-9 plays no direct functional role in HIF degradation since physical interaction of OS-9 with oxygen sensing HIF prolyl hydroxylases cannot occur in vivo due to their different subcellular localization

Oxygen Sensing in Drosophila: Multiple Isoforms of the Prolyl Hydroxylase Fatiga Have Different Capacity to Regulate HIFα/Sima

Background: The Hypoxia Inducible Factor (HIF) mediates cellular adaptations to low oxygen. Prolyl-4-hydroxylases are oxygen sensors that hydroxylate the HIF alpha-subunit, promoting its proteasomal degradation in normoxia. Three HIFprolyl hydroxylases, encoded by independent genes, PHD1, PHD2, and PHD3, occur in mammals. PHD2, the longest PHD isoform includes a MYND domain, whose biochemical function is unclear. PHD2 and PHD3 genes are induced in hypoxia to shut down HIF dependent transcription upon reoxygenation, while expression of PHD1 is oxygen-independent. The physiologic significance of the diversity of the PHD oxygen sensors is intriguing. Methodology and Principal Findings: We have analyzed the Drosophila PHD locus, fatiga, which encodes 3 isoforms, FgaA, FgaB and FgaC that are originated through a combination of alternative initiation of transcription and alternative splicing. FgaA includes a MYND domain and is homologous to PHD2, while FgaB and FgaC are shorter isoforms most similar to PHD3. Through a combination of genetic experiments in vivo and molecular analyses in cell culture, we show that fgaB but not fgaA is induced in hypoxia, in a Sima-dependent manner, through a HIF-Responsive Element localized in the first intron of fgaA. The regulatory capacity of FgaB is stronger than that of FgaA, as complete reversion of fga loss-of-function phenotypes is observed upon transgenic expression of the former, and only partial rescue occurs after expression of the latter. Conclusions and Significance: Diversity of PHD isoforms is a conserved feature in evolution. As in mammals, there are hypoxia-inducible and non-inducible Drosophila PHDs, and a fly isoform including a MYND domain co-exists with isoforms lacking this domain. Our results suggest that the isoform devoid of a MYND domain has stronger regulatory capacity than that including this domain.Fil:Acevedo, J.M. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Centanin, L. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Dekanty, A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Wappner, P. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina

Polybrene Inhibits Human Mesenchymal Stem Cell Proliferation during Lentiviral Transduction

Human mesenchymal stem cells (hMSCs) can be engineered to express specific genes, either for their use in cell-based therapies or to track them in vivo over long periods of time. To obtain long-term expression of these genes, a lentivirus- or retrovirus-mediated cell transduction is often used. However, given that the efficiency with these viruses is typically low in primary cells, additives such as polybrene are always used for efficient viral transduction. Unfortunately, as presented here, exposure to polybrene alone at commonly used concentratons (1–8 µg/mL) negatively impacts hMSC proliferation in a dose-dependent manner as measured by CyQUANT, EdU incorporation, and cell cycle analysis. This inhibition of proliferation was observable in culture even 3 weeks after exposure. Culturing the cells in the presence of FGF-2, a potent mitogen, did not abrogate this negative effect of polybrene. In fact, the normally sharp increase in hMSC proliferation that occurs during the first days of exposure to FGF-2 was absent at 4 µg/mL or higher concentrations of polybrene. Similarly, the effect of stimulating cell proliferation under simulated hypoxic conditions was also decreased when cells were exposed to polybrene, though overall proliferation rates were higher. The negative influence of polybrene was, however, reduced when the cells were exposed to polybrene for a shorter period of time (6 hr vs 24 hr). Thus, careful evaluation should be done when using polybrene to aid in lentiviral transduction of human MSCs or other primary cells, especially when cell number is critical

Nitrite-derived nitric oxide reduces hypoxia-inducible factor 1α-mediated extracellular vesicle production by endothelial cells

Introduction Extracellular vesicles (EVs) are small, spherical particles enclosed by a phospholipid bilayer (∼30–1000 nm) released from multiple cell types, and have been shown to have pathophysiological roles in a plethora of disease states. The transcription factor hypoxia-inducible factor-1 (HIF-1) allows for adaptation of cellular physiology in hypoxia and may permit the enhanced release of EVs under such conditions. Nitric oxide (NO) plays a pivotal role in vascular homeostasis, and can modulate the cellular response to hypoxia by preventing HIF-1 accumulation. We aimed to selectively target HIF-1 via sodium nitrite (NaNO2) addition, and examine the effect on endothelial EV, size, concentration and function, and delineate the role of HIF-1 in EV biogenesis. Methods Endothelial (HECV) cells were exposed to hypoxic conditions (1% O2, 24 h) and compared to endothelial cells exposed to normoxia (21% O2) with and without the presence of sodium nitrite (NaNO2) (30 μM). Allopurinol (100 μM), an inhibitor of xanthine oxidoreductase, was added both alone and in combination with NaNO2 to cells exposed to hypoxia. EV and cell preparations were quantified by nanoparticle tracking analysis and confirmed by electron microscopy. Western blotting and siRNA were used to confirm the role of HIF-1α and HIF-2α in EV biogenesis. Flow cytometry and time-resolved fluorescence were used to assess the surface and intravesicular protein content. Results Endothelial (HECV) cells exposed to hypoxia (1% O2) produced higher levels of EVs compared to cells exposed to normoxia. This increase was confirmed using the hypoxia-mimetic agent desferrioxamine. Treatment of cells with sodium nitrite (NaNO2) reduced the hypoxic enhancement of EV production. Treatment of cells with the xanthine oxidoreductase inhibitor allopurinol, in addition to NaNO2 attenuated the NaNO2-attributed suppression of hypoxia-mediated EV release. Transfection of cells with HIF-1α siRNA, but not HIF-2α siRNA, prior to hypoxic exposure prevented the enhancement of EV release. Conclusion These data provide evidence that hypoxia enhances the release of EVs in endothelial cells, and that this is mediated by HIF-1α, but not HIF-2α. Furthermore, the reduction of NO2− to NO via xanthine oxidoreductase during hypoxia appears to inhibit HIF-1α-mediated EV production